Comb-Like Fluorophilic-Lipophilic-Hydrophilic Polymers for Nanocapsules as Ultrasound Contrast Agents.

Imaging the enhanced permeation and retention effect by ultrasound is hindered by the large size of commercial ultrasound contrast agents (UCAs). To obtain nanosized UCAs, triblock copolymers of poly(ethylene glycol)-polylactide-poly(1 H,1 H,2 H,2 H-heptadecafluorodecyl methacrylate) (PEG-PLA-PFMA) with distinct numbers of perfluorinated pendant chains (5, 10, or 20) are synthesized by a combination of ring-opening polymerization and atom transfer radical polymerization. Nanocapsules (NCs) containing perfluorooctyl bromide (PFOB) intended as UCAs are obtained with a 2-fold increase in PFOB encapsulation efficiency in fluorinated NCs as compared with plain PEG-PLA NCs thanks to fluorous interactions. NC morphology is strongly influenced by the number of perfluorinated chains and the amount of polymer used for formulation, leading to peculiar capsules with several PFOB cores at high PEG-PLA-PFMA20 amount and single-cored NCs with a thinner shell at low fluorinated polymer amount, as confirmed by small-angle neutron scattering. Finally, fluorinated NCs yield higher in vitro ultrasound signal compared with PEG-PLA NCs, and no in vitro cytotoxicity is induced by fluorinated polymers and their degradation products. Our results highlight the benefit of adding comb-like fluorinated blocks in PEG-PLA polymers to modify the nanostructure and enhance the echogenicity of nanocapsules intended as UCAs.

Lymphatic Transport and Lymph Node Location of Microspheres Subcutaneously Injected in the Vicinity of Tumors in a Rabbit Model of Breast Cancer.

PURPOSE: To assess the lymphatic transport of microparticles of 100 nm, 1 µm and 10 µm subcutaneously injected into the breast area of healthy and tumor-bearing rabbits, and to analyze their location in lymph node (LN) in relation to malignant cells. METHODS: Female rabbits (n = 9) bearing a VX2 tumor in one thoracic mammary gland were subcutaneously injected at D15 with polystyrene fluorescent particles around the nipple, on the tumor and on the healthy sides. The tumor and the LN measured by ultrasound at D9, D15 and D20 were explanted at D20. The LN metastases were evaluated by cytokeratin staining. LN uptake of the particles was measured by quantifying the green fluorescence surface in hot spot regions of healthy and pathologic LN. RESULTS: All animals developed mammary tumors. Metastases were found in 39% of LN from the tumor side. LN invasion was significantly lower for the 10 µm group versus the 100 nm group (p < 0.0348). The fully invaded area of metastatic LN contained significantly less 100 nm and 1 µm particles compared to the low and non-invaded regions and to the healthy LN. In the invaded LN, the 1 µm MS occupied more surface than the 100 nm particles. CONCLUSIONS: 1 µm MS arrived numerously into the areas low-invaded and non-invaded by the tumoral cells of the pathologic LN, but they were very rare in the fully invaded regions. Compared to the 100 nm nanospheres, the 1 µm were better retained (20 times) into the sentinel LN, showing the advantage of micrometric particles for lymph-targeted chemotherapy when injected before complete invasion by metastases.

Laparoscopic subperitoneal injection of chemo-loaded particles lowers tumor growth on a rabbit model of peritoneal carcinomatosis.

The purpose of our study was to assess the effect of controlled-release chemotherapy on the growth and viability of peritoneal carcinomatosis treated by subperitoneal injection in a rabbit VX2 model. A model of peritoneal carcinomatosis was created by laparoscopic injection of VX2 tumor in the left and right broad ligaments of 12 White New Zealand rabbits. At day 12, each tumor was randomly treated with a peritumoral injection of 0.5 mL microspheres loaded with doxorubicin (DEM-DOX) or unloaded (DEM-BLAND). Seven days after treatment, tumor volume, tumor viability in histology, local tumor necrosis in contact with DEM, and doxorubicin concentration profile around the drug eluting microspheres (DEM) were measured. Tumor volume was significantly lower in the DEM-DOX group (3.6 ± 3.2 cm3) compared with the DEM-BLAND group (8.9 ± 5.4 cm3) (p = 0.0425). The percentage of viable tumor tissue was significantly lower in the DEM-DOX group (38% ± 17%) compared with the DEM-BLAND group (56% ± 20%) (p = 0.0202). Tissue necrosis was observed around all DEM-DOX up to a distance of 1.094 ± 0.852 mm and never observed around DEM-BLAND. Drug concentration was above the therapeutic level of 1.0 µM up to a distance of 1.4 mm from the DEM to the tumor. Laparoscopic subperitoneal injection of chemo-loaded particles is feasible and lowers tumor growth and viability in a rabbit model of peritoneal carcinomatosis after 1 week.

A novel resorbable embolization microsphere for transient uterine artery occlusion: a comparative study with trisacryl-gelatin microspheres in the sheep model.

PURPOSE: To evaluate angiographic recanalization, inflammatory reaction, and uterine damage after sheep uterine artery embolization (UAE) with a novel calibrated resorbable embolization microsphere (REM) and compare the results with control nonresorbable microspheres. MATERIALS AND METHODS: Six hormonally artificially cycled sheep underwent bilateral UAE until stasis with either REM or trisacryl-gelatin microspheres (TGMS). At 7 days, control angiograms were obtained to assess the residual vascularization at arterial and parenchymal phases. The animals were then sacrificed for analysis of the presence of microspheres, inflammatory foreign body reaction, and surface areas of uterine damage. RESULTS: Mean volume of microspheres injected per uterine artery (UA) or per animal did not differ between groups. At day 7, the flow was normal for six of six UAs that received embolization with REM versus only three of six UAs with TGMS (P = .0455, χ(2) test). Uterine parenchymography showed no defects in six UAs in the REM group versus five defects in six UAs in the TGMS group (P = .0060, χ(2) test). No REM or residual fragments of microspheres were observed on histologic analysis. TGMS were observed in tissues and accompanied by a mild inflammatory response. Necrosis rates were not significantly different between the two products, either in endometrium (REM 23.5% ± 28.8% [median 8.1%] vs TGMS 21.8% ± 23.7% [median 14.6%]) or in myometrium (REM 8.2% ± 22.7% [median 0.0%] vs TGMS 8.8% ± 20.8% [median 0.9%]). Endometrium alteration rate was lower with REM than with TGMS (39.7% ± 25.7% [median 34%] vs 60.6% ± 27.1% [median 71%]; P = .0060, Mann-Whitney test). Myometrium alteration rates were not significantly different between REM (45.7% ± 37.1% [median 63.0%]) and TGMS (37.8% ± 34.0% [median 19.1%]). CONCLUSIONS: At 1 week after sheep UAE with REM, the recanalization was complete, the microspheres were completely degraded, and there was no remnant inflammatory response.

Poly(malic acid)-budesonide nanoconjugates embedded in microparticles for lung administration.

To improve the therapeutic activity of inhaled glucocorticoids and reduce potential side effects, we designed a formulation combining the advantages of nanoparticles, which have an enhanced uptake by alveolar cells, allow targeted delivery and sustained drug release, as well as limited drug systemic passage, with those of microparticles, which display good alveolar deposition. Herein, a polymer-drug conjugate, poly(malic acid)-budesonide (PMAB), was first synthesized with either 11, 20, 33, or 43 mol% budesonide (drug:polymer from 1:8 to 3:4), the drug creating hydrophobic domains. The obtained conjugates self-assemble into nanoconjugates in water, yielding excellent drug loading of up to 73 wt%, with 80-100 nm diameters. In vitro assays showed that budesonide could be steadily released from the nanoconjugates, and the anti-inflammatory activity was preserved, as evidenced by reduced cytokine production in LPS-activated RAW 264.7 macrophages. Nanoconjugates were then embedded into microparticles through spray-drying with L-leucine, forming nano-embedded microparticles (NEMs). NEMs were produced with an aerodynamic diameter close to 1 µm and a density below 0.1 g.cm-3, indicative of a high alveolar deposition. NEMs spray-dried with the less hydrophobic nanoconjugates, PMAB 1:4, were readily dissolved in simulated lung fluid and were chosen for in vivo experiments to study pharmacokinetics in healthy rats. As it was released in vivo from NEMs, sustained distribution of budesonide was obtained for 48 h in lung tissue, cells, and lining fluid. With high loading rates, modulable release kinetics, and low cytotoxicity, these nanoconjugates delivered by NEMs are promising for the more efficient treatment of pulmonary inflammatory diseases.

Targeting and recanalization after embolization with calibrated resorbable microspheres versus hand-cut gelatin sponge particles in a porcine kidney model.

PURPOSE: To report on polyethylene glycol hydrogel-based resorbable embolization microspheres (REM) that were synthesized to resorb in < 24 hours, before inflammation and vascular remodeling, to achieve a complete arterial recanalization and to compare targeting and recanalization of REM of 300-500 µm, 500-700 µm, and 700-900 µm with hand-cut gelatin sponge particles (GSP). MATERIALS AND METHODS: Eight pigs underwent polar renal artery embolization with REM or GSP. Angiograms were obtained before embolization and 10 minutes and 7 days after embolization before pigs were sacrificed to determine the occlusion level, the percentage of occlusion, and the recanalization rate for each product. The distribution of embolic material was assessed in pathology, and infarction rate of the kidneys was measured. RESULTS: REM of 300-500 µm occluded more distal vessels than REM of 500-700 µm and 700-900 µm. At day 7, the recanalization rate was complete for the larger REM, whereas it was about 60% for the two smaller sizes. REM were completely degraded, with no residual material or inflammation. GSP occluded more proximal arteries than REM of 700-900 µm, were partly degraded at day 7, and were accompanied by a foreign body reaction in proximal and distal arteries. GSP recanalized at 79%. The infarction rate was higher with the two smaller sizes of REM and with GSP than with the largest REM. CONCLUSIONS: REM of different sizes targeted different occlusion levels in kidney arteries. GSP provided an extended occlusion level without actual targeting. Regardless of embolic material used, angiographic recanalization of renal arteries depended on the extent of necrosis. REM of 700-900 µm demonstrated the lowest infarction rate and the best recanalization rate.

Emerging nanoparticle platforms to improve the administration of glucocorticoids.

Glucocorticoids (GC) are common drugs used to treat acute and chronic inflammatory diseases, whose prolonged use can result in severe side effects hampering their efficacy. In addition, the pharmacokinetics, and biodistribution of GC are inadequate to support high efficacy with reduced toxicity. Following the marketing of GC prodrugs, new GC prodrug entities, and conjugates, have been developed. These new prodrugs and conjugates have been administered in free form or under a nanoparticulate form for local or systemic administration. These nanoparticles from lipid prodrugs and nanoconjugates change the paradigm of GC delivery, solving the issue of low drug loading into nanoparticles and circumventing the potential burst release effect by allowing a more controlled delivery of the GC and better targeting in inflammatory sites. This review highlights the design strategies, recent advances in GC prodrugs and conjugates, and their delivery in nanoparticulate form, demonstrating the strong potentialities of these novel strategies.

Novel surfactants with diglutamic acid polar head group: drug solubilization and toxicity studies.

PURPOSE: Novel surfactants made of diglutamic acid (DG) polar head linked to lithocholic, arachidonic, linoleic or stearic acids were designed for drug solubilization. METHODS: Surfactants 3-D conformer and packing parameter were determined by molecular modelling and self-assembling properties by pyrene fluorescence measurements. Cytotoxicity was assessed on Human Umbilical Vein Endothelial Cells (HUVEC) and haemolyitic activity on rat red blood cells. Drug solubilization was quantified and its interaction with hydrophobic moieties was characterized using differential scanning calorimetry and X-ray diffraction. Self organisation of stearoyl-DG was observed by cryogenic transmission electron microscopy. Toxicity after repeated injections of stearoyl-DG was investigated in Wistar rats. RESULTS: DG-based surfactants self-assemble into water and their critical micellar concentrations are comprised between 200 and 920 µg/mL. Cytotoxicity and haemolysis were lower than for polysorbate 80. At best, stearoyl-DG solubilized the drug up to 22% (w/w). Solid-state characterization evidenced drug/lipid interactions leading to the formation of a new complex. Stearoyl-DG formed spherical micelles of 20 nm, as predicted by packing parameter calculation. However, it induced a possible liver toxicity after intravenous administration in rats. CONCLUSIONS: Among the surfactants tested, stearoyl-DG is the more efficient for drug solubilization but its use is limited by its possible liver toxicity.

Mannosylation of budesonide palmitate nanoprodrugs for improved macrophage targeting.

In a strategy to improve macrophage targeting of glucocorticoids (GCs) for anti-inflammatory therapy, a so-called nanoprodrug of budesonide palmitate decorated by mannose moieties was designed. The synthesis of budesonide palmitate (BP) was obtained by esterification and mannosylated lipid (DSPE-PEG-Man) by reacting 1,2-Distearoyl-sn-Glycero-3-Phosphoethanolamine (DSPE)-polyethylene glycol-amine and α-D-mannopyranosylphenyl isothiocyanate (MPITC). Nanoparticles were formulated by emulsion-evaporation and different ratios of mannosylated lipid were introduced in the formulation of BP nanoprodrugs. Using up to 75% of DSPE-PEG-man (75/25) led to 200 nm particles with a polydispersity index below 0.2, a negative zeta potential ranging from -10 to -30 mV, and one-month stability at 4 °C. The encapsulation efficiency of BP approached 100% proving that the prodrug was associated with the particles, leading to a final BP loading of 50-to 60% (w/w). The lectin agglutination test confirmed the availability of mannose on the nanoprodrug surface. Nanoprodrug uptake by RAW 264.7 macrophages was observed by confocal microscopy and flow cytometry. After 24 and 48 h of incubation, a significantly greater internalization of mannosylated nanoparticles as compared to PEGylated nanoparticles was achieved. The mannose receptor-mediated uptake was confirmed by a mannan inhibition study. After LPS-induced inflammation, the anti-inflammatory effect of mannosylated nanoparticles was assessed. After 48 h of incubation, cytokines (MCP-1 and TNFα) were reduced demonstrating that the functionalization of nanoprodrugs is possible and efficient.

Biocompatible and Photostable Photoacoustic Contrast Agents as Nanoparticles Based on Bodipy Scaffold and Polylactide Polymers: Synthesis, Formulation, and In Vivo Evaluation.

We have designed a new Bodipy scaffold for efficient in vivo photoacoustic (PA) imaging of nanoparticles commonly used as drug nanovectors. The new dye has an optimized absorption band in the near-infrared window in biological tissue and a low fluorescence quantum yield that leads to a good photoacoustic generation efficiency. After Bodipy-initiated ring-opening polymerization of lactide, the polylactide-Bodipy was formulated into PEGylated nanoparticles (NPs) by mixing with PLA-PEG at different concentrations. Formulated NPs around 100 nm exhibit excellent PA properties: an absorption band at 760 nm and a molar absorption coefficient in between that of molecular PA absorbers and gold NPs. Highly improved photostability compared to cyanine-labeled PLA NPs as well as innocuity in cultured macrophages were demonstrated. After intravenous injection in healthy animals, NPs were easily detected using a commercial PA imaging system and spectral unmixing, opening the way to their use as theranostic agents.

Sustained-hepatic arterial infusion of oxaliplatin: pharmacokinetic advantages over hepatic arterial infusion using a preclinical animal tumour model.

Hepatic arterial infusion (HAI) of oxaliplatin allows greater liver tumour drug exposure compared to systemic infusion. However, the therapeutic index of HAI oxaliplatin remains poor. Using Pickering emulsion technology, we developed a platform able to provide sustained releases of oxaliplatin. The goal of this study was to evaluate the pharmacokinetic advantages of sustained-HAI oxaliplatin over HAI using a preclinical animal tumour model. Injections of 0.6 mg oxaliplatin in 20 min were selectively done in left hepatic arteries of 20 rabbits bearing a VX2 liver tumour in the middle left-lobe, using HAI (n = 10) or sustained-HAI (n = 10). In each group, half of the rabbits were sacrificed at 24 h and half at 72 h. Mass spectrometry was used to quantify drug pharmacokinetics in blood and oxaliplatin concentrations in tumour tissues, right- and middle left-liver lobes, spleen and lung. Compared to HAI, sustained-HAI of oxaliplatin resulted in lower plasmatic peak (Cmax: 275 ± 41 vs. 416 ± 133 ng/mL, p = 0.02) and higher concentration in the tumour at 24 h (2118 ± 2107 vs. 210 ± 93 ng/g, p = 0.008). After HAI, oxaliplatin concentration in tumours was significantly higher than in lung at 24 h (p = 0.03) but no other difference was found between oxaliplatin concentrations in tumours and in liver lobes, spleen or lung, neither at 24 h nor at 72 h. On the opposite, sustained-HAI resulted in higher concentrations of oxaliplatin in tumour compared to oxaliplatin concentrations in the middle left lobe (163 ± 86 ng/g at 24 h, p = 0.01, and 90 ± 15 ng/g at 72 h, p = 0.04), right lobe (174 ± 112 ng/g at 24 h, p = 0.01, and 112 ± 35 ng/g, p = 0.04 at 72 h), spleen (142 ± 21 ng/g at 24 h, p = 0.01, and 98 ± 12 ng/g at 72 h, p = 0.04), and lung (85 ± 11 ng/g at 24 h, p = 0.01, and 52 ± 4 ng/g at 72 h, p = 0.03). Sustained-HAI improves the therapeutic index of HAI oxaliplatin and offers a great potential for patients suffering from unresectable colorectal liver metastases or hepatocellular carcinoma.

Tuning morphology of Pickering emulsions stabilised by biodegradable PLGA nanoparticles: How PLGA characteristics influence emulsion properties.

In this study, we proved that the stabilisation of Pickering emulsions by polymer nanoparticles (NPs) heavily depends on polymer characteristics. We prepared NPs with four poly(lactide-co-glycolide) polymers (PLGA), of different molar masses (14,000 and 32,000 g/mol) and end groups (acid or alkylester). NPs were either bare (without stabilising polymer) or covered by polyvinyl alcohol (PVA). Pickering emulsions were prepared by mixing NP aqueous suspensions with various amounts of oil (Miglyol 812 N). First, NP wettability was directly affected by PLGA end group: ester-ending PLGA led to more hydrophobic NPs, compared to acid-ending PLGA. This effect of the end group could be slightly enhanced with smaller molar mass. Thus, bare PLGA NPs stabilised different types of emulsions (W/O/W and W/O), following Finkle's rule. However, the effect of PLGA characteristics was masked when NPs were covered by PVA, as PVA drove the stabilisation of O/W emulsions. Secondly, PLGA molar mass and end group also influenced its glass transition temperature (Tg), with spectacular consequences on emulsion formation. Indeed, the shortest ester-ending PLGA exhibited a Tg close to room temperature, when measured in the emulsion. This Tg, easily exceeded during emulsification process, led to a soft solid emulsion, stabilised by a network of NP debris.

Coadministration of nanosystems of short silencing RNAs targeting oestrogen receptor alpha and anti-oestrogen synergistically induces tumour growth inhibition in human breast cancer xenografts.

The suppression of oestrogen receptor alpha (ERalpha) functions by silencing RNAs in association with or not with anti-oestrogens (AEs) both in vitro and in breast cancer cell xenografts was assessed. In vitro, a prolonged decrease in ERalpha protein expression and an enhanced AE-induced inhibition of ERalpha-mediated transcription, together with antiproliferative activity, were observed. Incorporation of ERalpha-siRNAs in pegylated nanocapsules (NC) was achieved; and their intravenous injections in MCF-7 xenografts, in contrast to scramble siRNA containing NCs, lead to decrease in ERalpha protein content and Ki67 labelling in tumour cells. The pure AE RU58668 (RU) both free and entrapped in stealth nanospheres (NS) at very low concentration (8 microg/kg/week) had no effect on tumour growth evolution. However, coinjection of the two nanocarriers potentiated the decrease in ERalpha protein, concomitantly with decreasing tumour vasculature and glucose transporter-1. These data support that the targeted delivery of ERalpha-siRNA in breast tumours potentiates the inhibition of E(2)-induced proliferative activity by encapsulated AE through enhanced anti-vascular activity. In the hormone-independent MDA-MB-231 xenograft model, RU-NS at 4 mg/kg/week induce also a strong tumour vascular normalisation. Together, these findings suggest that the anti-oestrogen activity of RU as well as that of targeted ERalpha-siRNA leads to anti-angiogenic activity. Their delivery in stealth nanocarriers may constitute a new anti-cancer therapeutic strategy in solid tumours.

Pickering emulsions with ethiodized oil and nanoparticles for slow release of intratumoral anti-CTLA4 immune checkpoint antibodies.

BACKGROUND: Intratumorous immunotherapy for cancer is currently thriving. The aim of such local strategy is to improve the therapeutic index of these treatments, for higher on-target/on-tumor activity and less on-target/off-tumor adverse events. Strategies allowing for slow release of anti-CTLA4 in the tumor microenvironment could improve their clinical efficacy.The purpose of the study was to develop a radiopaque delivery platform to improve the targeting and exposure of intratumorous anti-CTLA4 antibodies for cancer immunotherapy. METHODS: Pickering emulsions of anti-CTLA4 antibodies were formulated with radiopaque ethiodized oil and poly-lactic-co-glycolic acid (PLGA) nanoparticles. We characterized the microscopic aspect and stability of such emulsions using Turbiscan. We monitored the release of anti-CTLA4 over time from these emulsions and evaluated their structure using mass spectrometry. We then tested the functionality of the released antibodies by preforming ex vivo competitive binding assays. Finally, we assessed the in vivo efficacy of intratumorous anti-CTLA4 Pickering emulsions. RESULTS: Pickering emulsions of ethiodized oil and PLGA nanoparticles (PEEPs) resulted in a radiopaque water-in-oil emulsion with average internal phase droplet size of 42±5 µm at day 7. Confocal microscopy showed that anti-CTLA4 antibodies were effectively encapsulated by ethiodized oil with PLGA nanoparticles located at the interface between the aqueous and the oily phase. Turbiscan analysis showed that emulsions were stable with continuous and progressive release of anti-CTLA4 antibodies reaching 70% at 3 weeks. Structural and functional analysis of the released antibodies did not show significant differences with native anti-CTLA4 antibodies. Finally, intratumorous anti-CTLA4 PEEPs were able to eradicate tumors and cure mice in a syngeneic immunocompetent preclinical tumor model. CONCLUSION: Pickering emulsions of ethiodized oil and PLGA is an innovative radiopaque delivery platform that does not alter the functionality of anti-CTLA4 immune checkpoint antibodies. Beyond local anti-CTLA4 applications, these emulsions might be used with other therapeutic molecules for optimal intratumorous or intra-arterial delivery of novel cancer immunotherapies.

Biodegradable Pickering emulsions of Lipiodol for liver trans-arterial chemo-embolization.

Water-in-oil (W/O) Lipiodol emulsions remain the preferable choice for local delivery of chemotherapy in the treatment of hepatocellular carcinoma. However, their low stability severely hampers their efficiency. Here, remarkably stable W/O Lipiodol emulsion stabilized by biodegradable particles was developed thanks to Pickering technology. The addition of poly(lactide-co-glycolide) nanoparticles (NPs) into the aqueous-phase of the formulation led to W/O Pickering emulsion by a simple emulsification process through two connected syringes. Influence of nanoparticles concentration and water/oil ratio on emulsion stability and droplet size were studied. All formulated Pickering emulsions were W/O type, stable for at least one month and water droplets size could be tuned by controlling nanoparticle concentration from 24 µm at 25 mg/mL to 69 µm at 5 mg/mL. The potential of these emulsions to efficiently encapsulate chemotherapy was studied through the internalization of doxorubicin (DOX) into the aqueous phase with a water/oil ratio of 1/3 as recommended by the medical community. Loaded-doxorubicin was released from conventional emulsion within a few hours whereas doxorubicin from stable Pickering emulsion took up to 10 days to be completely released. In addition, in vitro cell viability evaluations performed on the components of the emulsion and the Pickering emulsion have shown no significant toxicity up to relatively high concentrations of NPs (3 mg/mL) on two different cell lines: HUVEC and HepG2. STATEMENT OF SIGNIFICANCE: We present an original experimental research in the field of nanotechnology for biomedical applications. In particular, we have formulated, thanks to Pickering technology, a new therapeutic emulsion stabilized with biodegradable PLGA nanoparticles. As far as we know, this is the first therapeutic Pickering emulsion reported in the literature for hepatocellular carcinoma. Such a new emulsion allows to easily prepare a predictable and stable lipiodolized emulsion having all the required characteristics for optimum tumor uptake. As demonstrated throughout our manuscript, emulsions stabilized with these nanoparticles have the advantage of being biodegradable, biocompatible and less toxic compared to usual emulsions stabilized with synthetic surfactants. These findings demonstrate the plausibility of the use of Pickering emulsions for chemoembolization as a therapeutic agent in extended release formulations.

Physicochemical characteristics and preliminary in vivo biological evaluation of nanocapsules loaded with siRNA targeting estrogen receptor alpha.

Specific siRNAs that target estrogen receptor alpha (ERalpha) were encapsulated in nanocapsules (NCs). We produced small (approximately 100-200 nm) ERalpha-siRNA NCs with a water core by incorporating two mixed duplexes of specific ERalpha-siRNAs (ERalpha-mix-siRNA) into NCs. The encapsulation yield that was obtained with poly(iso-butylcyanoacrylate) (PIBCA) NCs was low, whereas no release of trapped siRNA was observed for poly(ethylene)glycol-poly(D,L-lactide-co-glycolide) (PEG-PLGA) NCs. High levels of ERalpha-siRNA incorporation into PEG-epsilon-caprolactone-malic acid (PEG-PCL/MA) NCs (3.3 microM in a polymer solution at 16 mg/mL) were observed (72% yield). No difference in size or zeta potential was observed between siRNA NCs that were based on PEG-PCL/MA and empty NCs. Fluorescence quenching assays confirmed the incorporation of siRNA into the NC core. A persistent loss of ERalpha (90% over 5 days) was observed in MCF-7 human breast cancer cells that were exposed to PEG-PCL/MA NCs that were loaded with ERalpha-siRNA. The intravenous injection of these NCs into estradiol-stimulated MCF-7 cell xenografts led to a significant decrease in tumor growth and a decrease in ERalpha expression in tumor cells. These data indicate that a novel strategy, based on ERalpha-siRNA delivery, could be developed for the treatment of hormone-dependent breast cancers.

Pickering-Emulsion for Liver Trans-Arterial Chemo-Embolization with Oxaliplatin.

PURPOSE: Biodegradable polylactic-co-glycolic acid (PLGA) nanoparticles can adsorb at the water/oil interface to stabilize the emulsion (forming Pickering-emulsion). The purpose of this study was to compare the release profiles of oxaliplatin from Pickering-emulsion and Lipiodol-emulsion. MATERIALS/METHODS: Pickering-emulsions and Lipiodol-emulsions were both formulated with oxaliplatin (5 mg/mL) and Lipiodol (water/oil ratio: 1/3). For Pickering-emulsion only, PLGA nanoparticles (15 mg/mL) were dissolved into oxaliplatin before formulation. In vitro release of oxaliplatin from both emulsions was evaluated. Then, oxaliplatin was selectively injected into left hepatic arteries of 18 rabbits bearing VX2 liver tumors using either 0.5 mL Pickering-emulsion (n = 10) or 0.5 mL Lipiodol-emulsion (n = 8). In each group, half of the rabbits were killed at 1 h and half at 24 h. Mass spectrometry was used to quantify drug pharmacokinetics in blood and resulting tissue (tumors, right, and left livers) oxaliplatin concentrations. RESULTS: Pickering-emulsion demonstrated a slow oxaliplatin release compared to Lipiodol-emulsion (1.5 ± 0.2 vs. 12.0 ± 6% at 1 h and 15.8 ± 3.0 vs. 85.3 ± 3.3% at 24 h) during in vitro comparison studies. For animal model studies, the plasmatic peak (Cmax) and the area under the curve (AUC) were significantly lower with Pickering-emulsion compared to Lipiodol-emulsion (Cmax = 0.49 ± 0.14 vs. 1.08 ± 0.41 ng/mL, p = 0.01 and AUC = 19.8 ± 5.9 vs. 31.8 ± 14.9, p = 0.03). This resulted in significantly lower oxaliplatin concentrations in tissues at 1 h with Pickering-emulsion but higher ratio between tumor and left liver at 24 h (43.4 vs. 14.5, p = 0.04). CONCLUSION: Slow release of oxaliplatin from Pickering-emulsion results in a significant decrease in systemic drug exposure and higher ratio between tumor and left liver oxaliplatin concentration at 24 h.

[The new biomaterials of embolization: the shift to degradable]. / Les nouveaux biomatériaux de l'embolisation - La grande révolution du dégradable.

During the last decade, the interest of degradable embolics has considerably grown as alternatives for the currently-used permanent embolics: no permanent foreign body, recanalisation for repeat embolization, complete drug delivery in chemoembolization. Their design is not trivial since a lot of requirements need to be satisfied. The degradable embolics should be easily suspended in physiological solutions and contrast media, injectable in catheters with small internal lumen and they have to recover their size and shape after injection, as for any embolic. Moreover, they need to be loadable with various drugs, and their degradation has to be achieved in a given time before the onset of a chronic inflammatory response and vessel wall remodeling. Various approaches have been tested on diverse materials over the last years and have produced interesting results that make it possible to claim that the revolution of the degradable embolic has begun.

Echogenicity enhancement by end-fluorinated polylactide perfluorohexane nanocapsules: Towards ultrasound-activable nanosystems.

Polylactide (PLA) polymers containing five distinct lengths of fluorinated (from C3F7 to C13F27) and non-fluorinated (C6H13) end-groups were successfully synthesized by ring-opening polymerization of d,l-lactide. Fluorination was expected to increase the encapsulation efficiency of perfluorohexane (PFH). 150 nm nanocapsules were obtained and 19F nuclear magnetic resonance revealed that nanocapsules formulated with fluorinated polymers increased by 2-fold the encapsulation efficiency of PFH compared with non-fluorinated derivatives, without any effect of fluorine chain length. Fluorination of the polymers did not induce any specific in vitro cytotoxicity of nanocapsules towards HUVEC and J774.A1 cell lines. The echogenicity of fluorinated-shelled nanocapsules was increased by 3-fold to 40-fold compared to non-fluorinated nanocapsules or nanoparticles devoid of a perfluorohexane core for both conventional and contrast-specific ultrasound imaging modalities. In particular, an enhanced echogenicity and contrast-specific response was observed as the fluorinated chain-length increased, probably due to an increase of density and promotion of bubble nucleation. When submitted to focused ultrasound, both intact and exploded nanocapsules could be observed, also with end-group dependency, indicating that PFH was partly vaporized. These results pave the way to the design of theranostic perfluorohexane nanocapsules co-encapsulating a drug for precision delivery using focused ultrasound. STATEMENT OF SIGNIFICANCE: We have synthesized novel fluorinated polyesters and formulated them into nanocapsules of perfluorohexane as ultrasound contrast agents. This nanosystem has been thoroughly characterized by several techniques and we show that fluorination of the biodegradable polymer favors the encapsulation of perfluorohexane without producing further reduction of cell viability. Contrary to nanocapsules of perfluoroctyl bromide formulated with the fluorinated polymers [32], the presence of the fluorinated moieties leads to an increase of echogenicity that is dependent of the length of the fluorinated moiety. Morevover, the ability of nanocapsules to explode when submitted to focused ultrasound also depends on the length of the fluorinated chain. These results pave the way to theranostic perfluorohexane nanocapsules co-encapsulating a drug for precision delivery using focused ultrasound.

Tunable delivery of niflumic acid from resorbable embolization microspheres for uterine fibroid embolization.

Uterine arteries embolization (UAE) is a recent technique that aims, by means of particles injected percutaneously, to stifle fibroids (leiomyomas). This treatment is non-invasive, compared with uterine ablation, but generates pelvic pain for a few days. A strategy to reduce the post-embolization pain would be to use calibrated embolization microspheres preloaded with a non-steroidal inflammatory drug (NSAID). In this study, we first compared four drugs, all active at low concentration on cyclooxygenase-2, i.e. ketoprofen, sodium diclofenac, flurbiprofen and niflumic acid (NFA), for their capacity to be loaded on resorbable embolization microspheres (REM) 500-700µm. NFA had the highest capacity of loading (5mg/mL) on resorbable microspheres. Then, we evaluated in vitro the NFA release profiles from REM having various degradation times of one, two or five days. NFA release was biphasic, with an initial burst (about 60% of the loading) followed by a sustained release that correlated significantly to REM's hydrolysis (rho=0.761, p<0.0001). For each group of beads, the size distribution was not modified by the loading of NFA and their delivery through microcatheter was not impaired by the drug. NFA eluted from REM inhibited the synthesis of prostaglandin E2 from rabbit uterus explants. In summary, NFA is loadable on REM in significant amount and its delivery can be tuned according to the degradation rate of REM to provide an antalgic effect for a few days after UAE.